NIR fluorescence-based imaging finds various applications in areas ranging from assessment of neuronal activity to sentinel lymph node identification and improved tumor detection and delineation. Because of the potential for high-sensitivity tumor detection in real-time, interest of the scientific community has been recently directed to fluorescence imaging systems operating mostly in the first NIR window (or NIR-1) for image-guided uses. Systems leveraging fluorescent dyes such Indocyanine green (ICG) and methylene blue (MB) include a FLARE™ system, a Fluobeam system, SPY, FDPM, and a Photodynamic Eye system, for example. Most of these utilize either a single image sensor (typically, a CCD) to capture visible (a reference image) and fluorescence images sequentially, or multiple cameras to form images in different spectral ranges simultaneously or sequentially.
For imaging systems that produce multiple images with the use of multiple cameras and/or sequential image acquisition, subsequent image registration is required (see Zhu, N. et al, “Dual Mode optical imaging system for fluorescence image-guided surgery”, Opt. Lett. 39(13); 3830-3932, 2014). To properly coordinate different spatial parameters of the multiple images, such image processing must take into account changes in angular coordinate(s), potential relative motion between the system and the subject, or both. Other types of imagers include imaging lenses that are configured for measurements of different emission spectra in absence of the visible reference window (Gray, D. C. et al., “Dual-mode laparoscopic fluorescence image-guided surgery using a single camera”, Biomed. Opt. Express, 3(8):1880-1890, 2012) and/or specialized CMOS sensors that can collect light via red-green-blue channel(s) (RGB) as well as a single channel in NIR-1 (Chen. Z et al., “Single camera imaging system for color and near-infrared fluorescence image guided surgery”, Biomed. Opt. Express, 5(8): 2791-2797, 2014).
Fluorescent imaging systems and methodologies possess well-recognized shortcomings limiting the operational capabilities of these systems. For example, some systems providing fused visible/fluorescence imagery with the use of a single, the only, image sensor, employ silicon-based detection units the sensitivity of which is limited to the visible and/or NIR-I spectral bands, in which case expensive spectral filters are additionally required to maintain the spectral purity of the passbands of the light source(s) and the detector (due to the fact that the spectral separation between the spectral band of the source of light and spectral band(s) within which the optical detection is carried out is very narrow). On the other hand, images acquired with the systems that employ two or more separate detectors to capture the radiation in the visible, NIR-I, and/or NIR-II spectral bands must be fused to create a combined image. To effectuate quality image fusion, such imaging systems must maintain precise optical alignment and positioning during the imaging procedure, which adversely impacts their size, weight, and cost.
Accordingly, there remains a need for a system and methodology overcoming deficiencies of the related art and enabling real-time detection and assessment of distribution of targets in object, both those located at or near the surface and those embedded within the object (such as, in a specific non-limiting case, the distribution of tumors in a tissue sample) for imaging of those targets.